Field of the Invention
The present invention relates generally to the analysis of chemical, biological, and cellular materials or structures and, more in particular, to a modified modular, portable and interchangeable analyte concentrator-microreactor (ACM) device, containing one or more immobilized affinity ligand groups, that can be coupled or mounted to a cartridge-cassette of a capillary electrophoresis apparatus or another similar unit of any other analytical separation apparatus. The analyte concentrator-microreactor (ACM) device is a functional and integral component of a cartridge-cassette of an analytical separation apparatus, to be used for the capture, isolation, purification, concentration, and release of simple and/or complex molecules, materials, and/or structures, as a whole or parts, to further be separated by a separation capillary or passage, followed by detection, quantification and characterization employing appropriate conditions for separation and the use of one or more detectors.
Description of Related Art
Microfabrication techniques have improved rapidly over the last decade, stimulated primarily by advancements in the microprocessor industry. Microfabrication in the field of analytical chemistry is having an impact on the manufacturing of small-sized analytical instrumentation and methods. Smaller-scale analysis often reduces requirements for reagents, electrical power and allows the manufacture of portable instruments that can be used in the field and in remote locations. Furthermore, microfabrication techniques facilitate the analysis of a sample in short periods of time, at high sensitivity, with high-throughput multi-dimensional/multi-task capabilities, environmental friendly, easy to operate and of interpreting the results, and being cost effective. Miniaturized platform technologies are one of the main challenges in medical device technology, in particular in the area of diagnostics. New and more versatile point-of-care diagnostic devices and methods are having an impact in the decentralization of diagnostics, helping to make disease diagnosis faster and more accurate, and facilitating decisions for an appropriate therapy and/or surgery if necessary. The increasing knowledge of the complex nature of molecular interactions has enabled not only a better understanding of physiological and pathological processes, but also the identification of biological markers (biomarkers) that define a particular state or condition.
Molecular and cellular biomarkers are now used across many disciplines and can be any molecule or cellular structure, part of a molecule or cellular structure, or even a particular configuration that is both detectable and measurable, where the amount, appearance or other property is indicative of a particular biological state. Typically diagnostic tests have been based on single biomarkers; however, basing a clinical decision on a single biomarker can lead to a significant level of false positives. As a consequence, multiplexing of biomarkers (e.g., signatures or panels) is being used to provide improved sensitivity, specificity and accuracy for the diagnosis and characterization of disease.
With the help of a panel of biomarkers it is possible to assess a disease at the very early stage of its formation, which is also helping in the understanding of how a selective treatment for a particular disease can work. The dogma is changing from using diagnostic tests mainly for diagnosing an advance stage of a disease, with defined symptoms, to a more predictive and pre-symptomatic analysis of a disease at an early stage of formation. There is a strong clinical imperative to identify discerning molecular biomarkers of disease that offer early detection of pathogenesis, inform prognosis, guide therapy, and monitor disease progression.
Therefore, there is an enduring need for improving instruments, technologies, and methodologies to identify biologically fluid-based, non-invasive molecular tests representing such selective panels of discerning biomarkers. One such a technology is the coupling of selective affinity-capture techniques with analytical separation techniques, as has been demonstrated with immunoaffinity capillary electrophoresis (IACE). In the IACE technology, the analyte concentrator-microreactor (ACM) device, a key component of an IACE instrument, can be used with a wide range of materials and chemistries to immobilized affinity capture substances of interest. In turn, the immobilized affinity ligands can selectively or non-selectively trap and isolate a single material or substance, or a panel of materials or substances (e.g., biomarkers), as a whole or parts, which can identify one or more diseases. The advantage of merging strategies for immunoassays and capillary electrophoresis is that now the two technologies complement the strengths of each other and work as one efficient technology. Furthermore, the technology of solid-phase capture and affinity purification have a wide variety of applications in research, clinical, pharmaceutical, forensic, environmental, food and beverage, various industrial areas, military and law enforcement institutions, terrorism (chemical or biological weapons), counterfeiting, and cultural heritage (authenticity of art work) laboratories.
Micro-scale miniaturization, represented by the IACE technology that uses a wide range of immobilized affinity ligand groups, brings major benefits to the enrichment of analytes found at low concentrations in simple and complex biological matrices. Furthermore, due to its high-resolution two-dimensionality platform, IACE has the capability of separating and quantifying intact substances and their respective modified counterparts, degradation products and/or metabolites.
The entire composition of the human body is very complex. It is not known precisely how many molecular and structural entities are, but it is known that they are many types of cellular, chemical and biochemical components, as a whole or parts and/or complexes from them, having different compositions, functions, sizes, shapes and also they vary by individual. Furthermore, the range of concentration of molecules in biological systems spans many orders of magnitude. Therefore, the use of miniaturized instrumentations and devices in the discovery of biomarkers has been a challenge for scientists. In particular, when the volumes to be used are small (e.g., microliters, nanoliters) and when the biomarkers to be analyzed are found in sub-nanomolar concentrations.
Capillary electrophoresis (CE) technology, in the conventional format and in the microchip format, has become a powerful tool employed in many laboratories in the search of important biomarker of diseases. However, the major deficiency of the CE technology for the isolation and quantification of biomarkers of interest is the limits of detection (LOD), which are constrained by the small dimension of the capillary and its reduced pathlength that hinders conventional optical detection methods such as ultraviolet detection. The steps of analyte purification and concentration for constituents present primarily in complex matrices still remains a bottleneck in the process of sample preparation.
Sample preparation is widely accepted as the most labor-intensive and error-prone part of the bioanalytical process. Sample preparation has been identified as being a bottleneck in total analysis samples, where complex matrices are used. It has become necessary then to develop sample preparation techniques to a new improved level. An integral, multi-task analyte concentrator-microreactor (ACM) device can be used for miniaturization and integration of sample preparation on-line when coupled to analytical separation instrumentation, in order to reduce laboratory workload and increase analytical performance. Furthermore, the ACM device is suitable for use with most existing applications by scaling the reagent and sample volumes, which can range from sub-microliter volumes to several milliliters.
Analyte concentrator-microreactor (ACM) devices have been developed for selective and non-selective molecular consolidations. These analyte concentrator-microreactor (ACM) devices, which are used on-line with a capillary tube or capillary channel, have been described in U.S. Pat. Nos. 5,202,010; 6,406,604; 7,329,388, 7,736,480; 7,811,436; 8,007,724; 8,007,725; 8,030,092; 8,182,746; 8,268,247; and 8,703,061 which are incorporated by references in this disclosure. U.S. Pat. No. 5,741,639 discloses the use of molecular recognition elements. U.S. Pat. No. 5,800,692 discloses the use of a pre-separation membrane for concentrating a sample. U.S. Pat. No. 7,407,568 discloses the use of sol-gel coating for on-line preconcentration in capillary electrophoresis. U.S. Pat. No. 7,828,948 discloses the use of preconcentration and separation of analytes in microchannels. U.S. Pat. No. 7,959,861 discloses the use of integrated affinity microcolumns and affinity capillary electrophoresis.